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1
Cox, Rachel T., Li-Mei Pai, Catherine Kirkpatrick, Joel Stein, and Mark Peifer. "Roles of the C Terminus of Armadillo in Wingless Signaling in Drosophila." Genetics 153, no. 1 (September 1, 1999): 319–32. http://dx.doi.org/10.1093/genetics/153.1.319.
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Abstract Drosophila melanogaster Armadillo and its vertebrate homolog β-catenin play multiple roles during development. Both are components of cell-cell adherens junctions and both transduce Wingless (Wg)/Wnt intercellular signals. The current model for Wingless signaling proposes that Armadillo binds the DNA-binding protein dTCF, forming a bipartite transcription factor that activates Wingless-responsive genes. In this model, Armadillo's C-terminal domain is proposed to serve an essential role as a transcriptional activation domain. In Xenopus, however, overexpression of C-terminally truncated β-catenin activates Wnt signaling, suggesting that the C-terminal domain might not be essential. We reexamined the function of Armadillo's C terminus in Wingless signaling. We found that C-terminally truncated mutant Armadillo has a deficit in Wg-signaling activity, even when corrected for reduced protein levels. However, we also found that Armadillo proteins lacking all or part of the C terminus retain some signaling ability if overexpressed, and that mutants lacking different portions of the C-terminal domain differ in their level of signaling ability. Finally, we found that the C terminus plays a role in Armadillo protein stability in response to Wingless signal and that the C-terminal domain can physically interact with the Arm repeat region. These data suggest that the C-terminal domain plays a complex role in Wingless signaling and that Armadillo recruits the transcriptional machinery via multiple contact sites, which act in an additive fashion.
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Hatzfeld, Mechthild, Christof Haffner, Katrin Schulze, and Ute Vinzens. "The Function of Plakophilin 1 in Desmosome Assembly and Actin Filament Organization." Journal of Cell Biology 149, no. 1 (April 3, 2000): 209–22. http://dx.doi.org/10.1083/jcb.149.1.209.
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Plakophilin 1, a member of the armadillo multigene family, is a protein with dual localization in the nucleus and in desmosomes. To elucidate its role in desmosome assembly and regulation, we have analyzed its localization and binding partners in vivo. When overexpressed in HaCaT keratinocytes, plakophilin 1 localized to the nucleus and to desmosomes, and dramatically enhanced the recruitment of desmosomal proteins to the plasma membrane. This effect was mediated by plakophilin 1's head domain, which interacted with desmoglein 1, desmoplakin, and keratins in the yeast two-hybrid system. Overexpression of the armadillo repeat domain induced a striking dominant negative phenotype with the formation of filopodia and long cellular protrusions, where plakophilin 1 colocalized with actin filaments. This phenotype was strictly dependent on a conserved motif in the center of the armadillo repeat domain. Our results demonstrate that plakophilin 1 contains two functionally distinct domains: the head domain, which could play a role in organizing the desmosomal plaque in suprabasal cells, and the armadillo repeat domain, which might be involved in regulating the dynamics of the actin cytoskeleton.
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Mills, Ryan D., Terrence D. Mulhern, Heung-Chin Cheng, and Janetta G. Culvenor. "Analysis of LRRK2 accessory repeat domains: prediction of repeat length, number and sites of Parkinson's disease mutations." Biochemical Society Transactions 40, no. 5 (September 19, 2012): 1086–89. http://dx.doi.org/10.1042/bst20120088.
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Various investigators have identified the major domain organization of LRRK2 (leucine-rich repeat kinase 2), which includes a GTPase ROC (Ras of complex proteins) domain followed by a COR (C-terminal of ROC) domain and a protein kinase domain. In addition, there are four domains composed of structural repeat motifs likely to be involved in regulation and localization of this complex protein. In the present paper, we report our bioinformatic analyses of the human LRRK2 amino acid sequence to predict the repeat size, number and likely boundaries for the armadillo repeat, ankyrin repeat, the leucine-rich repeat and WD40 repeat regions of LRRK2. Homology modelling using known protein structures with similar domains was used to predict structures, exposed residues and location of mutations for these repeat regions. We predict that the armadillo repeats, ankyrin repeats and leucine-rich repeats together form an extended N-terminal flexible ‘solenoid’-like structure composed of tandem repeat modules likely to be important in anchoring to the membrane and cytoskeletal structures as well as binding to other protein ligands. Near the C-terminus of LRRK2, the WD40 repeat region is predicted to form a closed propeller structure that is important for protein complex formation.
4
Fraser, Paul E., Gang Yu, Lyne Lévesque, Masaki Nishimura, Dun-Sheng Yang, Howard T. J. Mount, David Westaway, and Peter H. St George-Hyslop. "Presenilin function: connections to Alzheimer's disease and signal transduction." Biochemical Society Symposia 67 (February 1, 2001): 89–100. http://dx.doi.org/10.1042/bss0670089.
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Missense mutations in presenilin 1 (PS1) and presenilin 2 (PS2) are associated with early-onset familial Alzheimer's disease which displays an accelerated deposition of amyloid plaques and neurofibrillary tangles. Presenilins are multi-spanning transmembrane proteins which localize primarily to the endoplasmic reticulum and the Golgi compartments. We have previously demonstrated that PS1 exists as a high-molecular-mass complex that is likely to contain several functional ligands. Potential binding proteins were screened by the yeast two-hybrid system using the cytoplasmically orientated PS1 loop domain which was shown to interact strongly with members of the armadillo family of proteins, including ϐ-catenin, p0071 and a novel neuron-specific plakophilin-related armadillo protein (NPRAP). Armadillo proteins can have dual functions that encompass the stabilization of cellular junctions/synapses and the mediation of signal transduction pathways. Our observations suggest that PS1 may contribute to both aspects of armadillo-related pathways involving neurite outgrowth and nuclear translocation of ϐ-catenin upon activation of the wingless (Wnt) pathway. Alzheimer's disease (AD)-related presenilin mutations exhibit a dominant gain of aberrant function resulting in the prevention of ϐ-catenin translocation following Wnt signalling. These findings indicate a functional role for PS1 in signalling and suggest that mistrafficking of selected presenilin ligands may be a potential mechanism in the genesis of AD.
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Fleckenstein, D., M. Rohde, D. J. Klionsky, and M. Rudiger. "Yel013p (Vac8p), an armadillo repeat protein related to plakoglobin and importin alpha is associated with the yeast vacuole membrane." Journal of Cell Science 111, no. 20 (October 15, 1998): 3109–18. http://dx.doi.org/10.1242/jcs.111.20.3109.
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Proteins of the armadillo family are involved in diverse cellular processes in higher eukaryotes. Some of them, like armadillo, beta-catenin and plakoglobins have dual functions in intercellular junctions and signalling cascades. Others, belonging to the importin-alpha-subfamily are involved in NLS recognition and nuclear transport, while some members of the armadillo family have as yet unknown functions. Here, we introduce the Saccharomyces cerevisiae protein Yel013p as a novel armadillo (arm) repeat protein. The ORF Yel013w was identified in the genome project on chromosome V (EMBL: U18530) and codes for an acidic protein of 578 residues with 8 central arm-repeats, which are closely related to the central repeat-domain of Xenopus laevis plakoglobin. We show that Yel013p (Vac8p) is constitutively expressed in diploid and haploid yeasts and that it is not essential for viability and growth. However, the vacuoles of mutant cells are multilobular or even fragmented into small vesicles and the processing of aminopeptidase I, representing the cytoplasm-to-vacuole transport pathway, is strongly impaired. Consistent with these observations, subcellular fractionation experiments, immunolocalization and expression of green fluorescent protein (GFP) fusion proteins revealed that Yel013p (Vac8p) is associated with the vacuolar membrane. Our data provide evidence for the involvement of an arm-family member in vacuolar morphology and protein targeting to the vacuole.
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Cavalcante, Isadora Pontes, Anna Vaczlavik, Ludivine Drougat, Claudimara Ferini Pacicco Lotfi, Karine Perlemoine, Christopher Ribes, Marthe Rizk-Rabin, et al. "Cullin 3 targets the tumor suppressor gene ARMC5 for ubiquitination and degradation." Endocrine-Related Cancer 27, no. 4 (April 2020): 221–30. http://dx.doi.org/10.1530/erc-19-0502.
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ARMC5 (Armadillo repeat containing 5 gene) was identified as a new tumor suppressor gene responsible for hereditary adrenocortical tumors and meningiomas. ARMC5 is ubiquitously expressed and encodes a protein which contains a N-terminal Armadillo repeat domain and a C-terminal BTB (Bric-a-Brac, Tramtrack and Broad-complex) domain, both docking platforms for numerous proteins. At present, expression regulation and mechanisms of action of ARMC5 are almost unknown. In this study, we showed that ARMC5 interacts with CUL3 requiring its BTB domain. This interaction leads to ARMC5 ubiquitination and further degradation by the proteasome. ARMC5 alters cell cycle (G1/S phases and cyclin E accumulation) and this effect is blocked by CUL3. Moreover, missense mutants in the BTB domain of ARMC5, identified in patients with multiple adrenocortical tumors, are neither able to interact and be degraded by CUL3/proteasome nor alter cell cycle. These data show a new mechanism of regulation of the ARMC5 protein and open new perspectives in the understanding of its tumor suppressor activity.
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Hansen, Simon, Christian Reichen, Chaithanya Madhurantakam, Markus Grütter, Andreas Plückthun, and Peer Mittl. "Structure-based engineering of designed armadillo repeat proteins." Acta Crystallographica Section A Foundations and Advances 70, a1 (August 5, 2014): C1153. http://dx.doi.org/10.1107/s2053273314088469.
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The specific recognition of macromolecules is key for many applications in biochemical research, medical diagnostics and disease treatment. Currently the development of new recognition molecules depends on the immunization of lab animals or combinatorial biochemistry techniques. Since both approaches are elaborate and require the availability of sufficient amounts of stable target molecules we are developing a modular system that allows a rational design of peptide recognition modules. This system is based on the armadillo repeat scaffold, because natural armadillo repeat proteins bind their targets in extended anti-parallel conformations with very regular binding topologies. The development of designed armadillo repeat proteins (dArmRPs) consisting entirely of identical internal repeats (full-consensus design) has been described [1]. Although dArmRP with 2nd generation capping repeats were predominantly monomeric in solution, crystal structures of dArmRP with different numbers of internal repeats revealed domain-swapped N-caps [2]. Redesign of the N-cap significantly improved thermodynamic stability and abrogated swapping of N-caps. These 3rd generation dArmRP recognize full-consensus peptides with nanomolar affinities. The dissociation constants depend on the lengths of the targeted peptides and the number of internal repeats. Three crystal structures of complexes between dArmRPs with five or six internal repeats and the targeted full-consensus (KR)5 peptide (either free or fused to the N- and C-terminii of globular proteins) confirm that the binding mode fulfills the expected regular topology. Further crystal structures of complexes between dArmRPs and the mismatch (RR)5 peptide revealed that the dArmRPs recognize their target peptides in a side-chain specific manner. Several crystal structures confirm that 3rd generation dArmRPs behave as stable and monomeric molecules that allow the selective recognition of the targeted peptide with the expected topology. Therefore, the rational assembly of binding modules from a pool of dipeptide-specific armadillo repeats to recognize peptides with given sequences should indeed be possible.
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Keil, René, Jenny Schulz, and Mechthild Hatzfeld. "p0071/PKP4, a multifunctional protein coordinating cell adhesion with cytoskeletal organization." Biological Chemistry 394, no. 8 (August 1, 2013): 1005–17. http://dx.doi.org/10.1515/hsz-2013-0114.
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Abstract P0071 is a member of a subfamily of armadillo proteins that also comprises p120-catenin (p120ctn), δ-catenin/NPRAP, ARVCF and the more distantly related plakophilins 1–3. These proteins share a conserved central domain consisting of a series of repeated motifs, the armadillo repeats, which is flanked by more diverse amino- and carboxy-terminal domains. P0071 and the related proteins were first described as components of adherens junctions with a function in clustering and stabilizing cadherins, thereby controlling intercellular adhesion. In addition, these proteins show a cytoplasmic and a nuclear localization. Major progress in understanding their cytoplasmic role has been made in recent years. One common theme appears to be the spatiotemporal control of the small GTPases of the Rho family in various cellular contexts, such as cell adhesion and motility, cell division or neurite outgrowth. In this review article, we focus on the functions of the p0071 protein and its closest relatives in regulating cell adhesion and cytoskeletal organization, which are critically involved in the control of cell polarity. Understanding p0071’s multiple functions requires assigning specific functions to particular binding partners and subcellular compartments. The identification of several new p0071 interacting proteins has promoted our understanding of the complex functions of this protein. Moreover, an initial analysis of its regulation begins to shed light on how these functions are coordinated in a cellular context.
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Palka, H. L., and K. J. Green. "Roles of plakoglobin end domains in desmosome assembly." Journal of Cell Science 110, no. 19 (October 1, 1997): 2359–71. http://dx.doi.org/10.1242/jcs.110.19.2359.
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Plakoglobin, a member of the armadillo family of proteins, is a component of intercellular adhesive junctions. The central domain of plakoglobin comprises a highly conserved series of armadillo repeats that facilitate its association with either desmosomal or classic cadherins, or with cytosolic proteins such as the tumor suppressor gene product adenomatous polyposis coli. Sequences in the N- and C-terminal domains of plakoglobin are less highly conserved, and their possible roles in regulating plakoglobin's subcellular distribution and junction assembly are still unclear. Here we have examined the role of plakoglobin end domains by stably expressing constructs lacking the N and/or C terminus of plakoglobin in A-431 cells. Our results demonstrate that myc-tagged plakoglobin lacking either end domain is still able to associate with the desmosomal cadherin desmoglein and incorporate into desmosomes. In cell lines that express an N-terminal truncation of plakoglobin, an increase in the cytosolic pool of en-dogenous and ectopic plakoglobin was observed that may reflect an increase in the stability of the protein. Deletion of the N terminus did not have a dramatic effect on the structure of desmosomes in these cells. On the other hand, striking alterations in desmosome morphology were observed in cells expressing C-terminal truncations of plakoglobin. In these cell lines, ectopic plakoglobin incorporated into desmosomes, and extremely long junctions or groups of tandemly linked desmosomes which remained well attached to keratin intermediate filaments, were observed. Together, these results suggest that plakoglobin end domains play a role in regulating its subcellular distribution, and that the presence of the C terminus limits the size of desmosomes, perhaps through regulating protein-protein interactions required for assembly of the desmosomal plaque.
10
Mariner, D. J., J. Wang, and A. B. Reynolds. "ARVCF localizes to the nucleus and adherens junction and is mutually exclusive with p120(ctn) in E-cadherin complexes." Journal of Cell Science 113, no. 8 (April 15, 2000): 1481–90. http://dx.doi.org/10.1242/jcs.113.8.1481.
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ARVCF is a novel Armadillo repeat domain protein that is closely related to the catenin p120(ctn). Using new ARVCF monoclonal antibodies, we have found that ARVCF associates with E-cadherin and competes with p120 for interaction with the E-cadherin juxtamembrane domain. ARVCF also localized to the nucleus in some cell types, however, and was significantly more nucleophilic than p120. Surprisingly, despite apparently ubiquitous expression, ARVCF was at least tenfold less abundant than p120 in a wide variety of cell types, and was difficult to detect by immunofluorescence unless overexpressed. Consequently, it is not likely to be abundant enough in adult tissues to functionally compete with p120. ARVCF also completely lacked the ability to induce the cell-branching phenotype associated with overexpression of p120. Expression of ARVCF/p120 chimeras confirmed previous results indicating that the branching activity of p120 maps to its Armadillo repeat domain. Surprisingly, the preferential localization of ARVCF to the nucleus required sequences in the amino-terminal end of ARVCF, suggesting that the sequences directing nuclear translocation of ARVCF are distinct from the predicted bipartite nuclear localization signal located between repeats 6 and 7. The dual localization of ARVCF to junctions and to nuclei suggests activities in different cellular compartments, as is the case for several other Armadillo repeat proteins including beta-catenin, p120 and the plakophilins.
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Barth, Angela I. M., Kathleen A. Siemers, and W. James Nelson. "Dissecting interactions between EB1, microtubules and APC in cortical clusters at the plasma membrane." Journal of Cell Science 115, no. 8 (April 15, 2002): 1583–90. http://dx.doi.org/10.1242/jcs.115.8.1583.
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End-binding protein (EB) 1 binds to the C-terminus of adenomatous polyposis coli (APC) protein and to the plus ends of microtubules (MT) and has been implicated in the regulation of APC accumulation in cortical clusters at the tip of extending membranes. We investigated which APC domains are involved in cluster localization and whether binding to EB1 or MTs is essential for APC cluster localization. Armadillo repeats of APC that lack EB1- and MT-binding domains are necessary and sufficient for APC localization in cortical clusters; an APC fragment lacking the armadillo repeats, but containing MT-and EB1-binding domains, does not localize to the cortical clusters but instead co-aligns with MTs throughout the cell. Significantly, analysis of endogenous proteins reveals that EB1 does not accumulate in the APC clusters. However, overexpressed EB1 does accumulate in APC clusters; the APC-binding domain in EB1 is located in the C-terminal region of EB1 between amino acids 134 and 268. Overexpressed APC- or MT-binding domains of EB1 localize to APC cortical clusters and MT, respectively, without affecting APC cluster formation itself. These results show that localization of APC in cortical clusters is different from that of EB1 at MT plus ends and appears to be independent of EB1.
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Froissard, Marine, Anne-Marie Keller, and Jean Cohen. "ND9P, a Novel Protein With Armadillo-like Repeats Involved in Exocytosis: Physiological Studies Using Allelic Mutants in Paramecium." Genetics 157, no. 2 (February 1, 2001): 611–20. http://dx.doi.org/10.1093/genetics/157.2.611.
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Abstract In Paramecium, a number of mutants affected in the exocytotic membrane fusion step of the regulated secretory pathway have been obtained. Here, we report the isolation of one of the corresponding genes, ND9, previously suspected to encode a soluble protein interacting with both plasma and trichocyst membranes. Nd9p is a novel polypeptide that contains C-terminal Armadillo-like repeats. Point mutations were found in the first N-terminal quarter of the molecule and in the last putative Armadillo repeat, respectively, for the two thermosensitive mutants, nd9-1 and nd9-2. The different behaviors of these mutants in recovery experiments upon temperature shifts suggest that the N-terminal domain of the molecule may be involved in membrane binding activity, whereas the C-terminal domain is a candidate for protein-protein interactions. The nonsense nd9-3 mutation that produces a short N-terminal peptide has a dominant negative effect on the nd9-1 allele. We show here that, when overexpressed, the dominant negative effect can be produced even on the wild-type allele, suggesting competition for a common target. We suggest that Nd9p could act, like some SNARE proteins, at the membrane-cytosol interface to promote membrane fusion.
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Knop, Jan, Tim Lienemann, Haifa El-Kilani, Sven Falke, Catharina Krings, Maria Sindalovskaya, Johannes Bergler, Christian Betzel, and Stefan Hoth. "Structural Features of a Full-Length Ubiquitin Ligase Responsible for the Formation of Patches at the Plasma Membrane." International Journal of Molecular Sciences 22, no. 17 (August 31, 2021): 9455. http://dx.doi.org/10.3390/ijms22179455.
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Plant U-box armadillo repeat (PUB-ARM) ubiquitin (Ub) ligases have important functions in plant defense through the ubiquitination of target proteins. Defense against pathogens involves vesicle trafficking and the formation of extracellular vesicles. The PUB-ARM protein SENESCENCE ASSOCIATED UBIQUITIN E3 LIGASE1 (SAUL1) can form patches at the plasma membrane related to tethering multi-vesicular bodies (MVBs) to the plasma membrane. We uncovered the structure of a full-length plant ubiquitin ligase and the structural requirements of SAUL1, which are crucial for its function in patch formation. We resolved the structure of SAUL1 monomers by small-angle X-ray scattering (SAXS). The SAUL1 model showed that SAUL1 consists of two domains: a domain containing the N-terminal U-box and armadillo (ARM) repeats and the C-terminal ARM repeat domain, which includes a positively charged groove. We showed that all C-terminal ARM repeats are essential for patch formation and that this function requires arginine residue at position 736. By applying SAXS to polydisperse SAUL1 systems, the oligomerization of SAUL1 is detectable, with SAUL1 tetramers being the most prominent oligomers at higher concentrations. The oligomerization domain consists of the N-terminal U-box and some N-terminal ARM repeats. Deleting the U-box resulted in the promotion of the SAUL1 tethering function. Our findings indicate that structural changes in SAUL1 may be fundamental to its function in forming patches at the plasma membrane.
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Wahl, J. K., J. E. Nieset, P. A. Sacco-Bubulya, T. M. Sadler, K. R. Johnson, and M. J. Wheelock. "The amino- and carboxyl-terminal tails of (beta)-catenin reduce its affinity for desmoglein 2." Journal of Cell Science 113, no. 10 (May 15, 2000): 1737–45. http://dx.doi.org/10.1242/jcs.113.10.1737.
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beta-catenin and plakoglobin are members of the armadillo family of proteins and were first identified as components of intercellular adhering junctions. In the adherens junction beta-catenin and plakoglobin serve to link classical cadherins to the actin-based cytoskeleton. In the desmosome plakoglobin links the desmosomal cadherins, the desmogleins and the desmocollins, to the intermediate filament cytoskeleton. beta-catenin is not a component of the desmosome. Previously we have shown that the central armadillo repeat region of plakoglobin is the site for desmosomal cadherin binding. We hypothesized that the unique amino- and/or carboxyl-terminal ends of beta-catenin may regulate its exclusion from the desmosomal plaque. To test this hypothesis we used chimeras between beta-catenin and plakoglobin to identify domain(s) that modulate association with desmoglein 2. Chimeric constructs, each capable of associating with classical cadherins, were assayed for association with the desmosomal cadherin desmoglein 2. Addition of either the N- or C-terminal tail of beta-catenin to the armadillo repeats of plakoglobin did not interfere with desmoglein 2 association. However, when both beta-catenin amino terminus and carboxyl terminus were added to the plakoglobin armadillo repeats, association with desmoglein 2 was diminished. Removal of the first 26 amino acids from this construct restored association. We show evidence for direct protein-protein interactions between the amino- and carboxyl-terminal tails of beta-catenin and propose that a sequence in the first 26 amino acids of beta-catenin along with its carboxyl-terminal tail decrease its affinity for desmoglein and prevent its inclusion in the desmosome.
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Tao, Y. S., R. A. Edwards, B. Tubb, S. Wang, J. Bryan, and P. D. McCrea. "beta-Catenin associates with the actin-bundling protein fascin in a noncadherin complex." Journal of Cell Biology 134, no. 5 (September 1, 1996): 1271–81. http://dx.doi.org/10.1083/jcb.134.5.1271.
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Catenins were first characterized as linking the cytoplasmic domains of cadherin cell-cell adhesion molecules to the cortical actin cytoskeleton. In addition to their essential role in modulating cadherin adhesivity, catenins have more recently been indicated to participate in cell and developmental signaling pathways. beta-Catenin, for example, associates directly with at least two receptor tyrosine kinases and transduces developmental signals within the Wnt pathway. Catenins also complex with the tumor suppressor protein adenomatous polyposis coli (APC), which appears to have a role in regulating cell proliferation. We have used the yeast two-hybrid method to reveal that fascin, a bundler of actin filaments, binds to beta-catenin's central Armadillo repeat domain. Western blotting of immunoprecipitates from cell line and mouse and rat brain extracts indicate that this interaction exists in vivo. Fascin and beta-catenin's association was further substantiated in vitro using purified proteins isolated from recombinant bacterial and baculoviral sources. Immunoprecipitation analysis indicates that fascin additionally binds to plakoglobin, which is highly homologous to beta-catenin but not to p120cas, a newly described catenin which contains a more divergent Armadillo-repeat domain. Immunoprecipitation, in vitro competition, and domain-mapping experiments demonstrate that fascin and E-cadherin utilize a similar binding site within beta-catenin, such that they form mutually exclusive complexes with beta-catenin. Immunofluorescence microscopy reveals that fascin and beta-catenin colocalize at cell-cell borders and dynamic cell leading edges of epithelial and endothelial cells. In addition to cell-cell borders, cadherins were unexpectedly observed to colocalize with fascin and beta-catenin at cell leading edges. It is conceivable that beta-catenin participates in modulating cytoskeletal dynamics in association with the microfilament-bundling protein fascin, perhaps in a coordinate manner with its functions in cadherin and APC complexes.
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Vincent, Peter A., Kanyan Xiao, Kathleen M. Buckley, and Andrew P. Kowalczyk. "VE-cadherin: adhesion at arm's length." American Journal of Physiology-Cell Physiology 286, no. 5 (May 2004): C987—C997. http://dx.doi.org/10.1152/ajpcell.00522.2003.
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VE-cadherin was first identified in the early 1990s and quickly emerged as an important endothelial cell adhesion molecule. The past decade of research has revealed key roles for VE-cadherin in vascular permeability and in the morphogenic events associated with vascular remodeling. The details of how VE-cadherin functions in adhesion became apparent with structure-function analysis of the cadherin extracellular domain and with the identification of the catenins, a series of cytoplasmic proteins that bind to the cadherin tail and mediate interactions between cadherins and the cytoskeleton. Whereas early work focused on the armadillo family proteins β-catenin and plakoglobin, more recent investigations have identified p120-catenin (p120 ctn) and a related group of armadillo family members as key binding partners for the cadherin tail. Furthermore, a series of new studies indicate a key role for p120 ctn in regulating cadherin membrane trafficking in mammalian cells. These recent studies place p120 ctn at the hub of a cadherin-catenin regulatory mechanism that controls cadherin plasma membrane levels in cells of both epithelial and endothelial origin.
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Daniel, Juliet M., and Albert B. Reynolds. "The Catenin p120ctnInteracts with Kaiso, a Novel BTB/POZ Domain Zinc Finger Transcription Factor." Molecular and Cellular Biology 19, no. 5 (May 1, 1999): 3614–23. http://dx.doi.org/10.1128/mcb.19.5.3614.
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ABSTRACT p120 ctn is an Armadillo repeat domain protein with structural similarity to the cell adhesion cofactors β-catenin and plakoglobin. All three proteins interact directly with the cytoplasmic domain of the transmembrane cell adhesion molecule E-cadherin; β-catenin and plakoglobin bind a carboxy-terminal region in a mutually exclusive manner, while p120 binds the juxtamembrane region. Unlike β-catenin and plakoglobin, p120 does not interact with α-catenin, the tumor suppressor adenomatous polyposis coli (APC), or the transcription factor Lef-1, suggesting that it has unique binding partners and plays a distinct role in the cadherin-catenin complex. Using p120 as bait, we conducted a yeast two-hybrid screen and identified a novel transcription factor which we named Kaiso. Kaiso’s deduced amino acid sequence revealed an amino-terminal BTB/POZ protein-protein interaction domain and three carboxy-terminal zinc fingers of the C2H2 DNA-binding type. Kaiso thus belongs to a rapidly growing family of POZ-ZF transcription factors that include the Drosophila developmental regulators Tramtrak and Bric à brac, and the human oncoproteins BCL-6 and PLZF, which are causally linked to non-Hodgkins’ lymphoma and acute promyelocytic leukemia, respectively. Monoclonal antibodies to Kaiso were generated and used to immunolocalize the protein and confirm the specificity of the p120-Kaiso interaction in mammalian cells. Kaiso specifically coprecipitated with a variety of p120-specific monoclonal antibodies but not with antibodies to α- or β-catenin, E-cadherin, or APC. Like other POZ-ZF proteins, Kaiso localized to the nucleus and was associated with specific nuclear dots. Yeast two-hybrid interaction assays mapped the binding domains to Arm repeats 1 to 7 of p120 and the carboxy-terminal 200 amino acids of Kaiso. In addition, Kaiso homodimerized via its POZ domain but it did not heterodimerize with BCL-6, which heterodimerizes with PLZF. The involvement of POZ-ZF proteins in development and cancer makes Kaiso an interesting candidate for a downstream effector of cadherin and/or p120 signaling.
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Tanentzapf, Guy, Christian Smith, Jane McGlade, and Ulrich Tepass. "Apical, Lateral, and Basal Polarization Cues Contribute to the Development of the Follicular Epithelium during Drosophila Oogenesis." Journal of Cell Biology 151, no. 4 (November 13, 2000): 891–904. http://dx.doi.org/10.1083/jcb.151.4.891.
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Analysis of the mechanisms that control epithelial polarization has revealed that cues for polarization are mediated by transmembrane proteins that operate at the apical, lateral, or basal surface of epithelial cells. Whereas for any given epithelial cell type only one or two polarization systems have been identified to date, we report here that the follicular epithelium in Drosophila ovaries uses three different polarization mechanisms, each operating at one of the three main epithelial surface domains. The follicular epithelium arises through a mesenchymal–epithelial transition. Contact with the basement membrane provides an initial polarization cue that leads to the formation of a basal membrane domain. Moreover, we use mosaic analysis to show that Crumbs (Crb) is required for the formation and maintenance of the follicular epithelium. Crb localizes to the apical membrane of follicle cells that is in contact with germline cells. Contact to the germline is required for the accumulation of Crb in follicle cells. Discs Lost (Dlt), a cytoplasmic PDZ domain protein that was shown to interact with the cytoplasmic tail of Crb, overlaps precisely in its distribution with Crb, as shown by immunoelectron microscopy. Crb localization depends on Dlt, whereas Dlt uses Crb-dependent and -independent mechanisms for apical targeting. Finally, we show that the cadherin–catenin complex is not required for the formation of the follicular epithelium, but only for its maintenance. Loss of cadherin-based adherens junctions caused by armadillo (β-catenin) mutations results in a disruption of the lateral spectrin and actin cytoskeleton. Also Crb and the apical spectrin cytoskeleton are lost from armadillo mutant follicle cells. Together with previous data showing that Crb is required for the formation of a zonula adherens, these findings indicate a mutual dependency of apical and lateral polarization mechanisms.
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Kausalya, P. Jaya, Dominic C. Y. Phua, and Walter Hunziker. "Association of ARVCF with Zonula Occludens (ZO)-1 and ZO-2: Binding to PDZ-Domain Proteins and Cell-Cell Adhesion Regulate Plasma Membrane and Nuclear Localization of ARVCF." Molecular Biology of the Cell 15, no. 12 (December 2004): 5503–15. http://dx.doi.org/10.1091/mbc.e04-04-0350.
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ARVCF, an armadillo-repeat protein of the p120ctnfamily, associates with classical cadherins and is present in adherens junctions, but its function is poorly understood. Here, we show that ARVCF interacts via a C-terminal PDZ-binding motif with zonula occludens (ZO)-1 and ZO-2. ARVCF and ZO-1 partially colocalize in the vicinity of the apical adhesion complex in polarized epithelial Madin-Darby canine kidney cells. ARVCF, ZO-1, and E-cadherin form a complex and are recruited to sites of initial cell-cell contact in sparse cell cultures. E-cadherin binding and plasma membrane localization of ARVCF require the PDZ-binding motif. Disruption of cell-cell adhesion releases ARVCF from the plasma membrane and an increased fraction of the protein localizes to the nucleus. Nuclear localization of ARVCF also requires the PDZ-binding motif and can be mediated by the PDZ domains of ZO-2. Thus, the interaction of ARVCF with distinct PDZ-domain proteins determines its subcellular localization. Interactions with ZO-1 and ZO-2, in particular, may mediate recruitment of ARVCF to the plasma membrane and the nucleus, respectively, possibly in response to cell-cell adhesion cues.
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Venkiteswaran, Kala, Kanyan Xiao, Susan Summers, Cathárine C. Calkins, Peter A. Vincent, Kevin Pumiglia, and Andrew P. Kowalczyk. "Regulation of endothelial barrier function and growth by VE-cadherin, plakoglobin, and β-catenin." American Journal of Physiology-Cell Physiology 283, no. 3 (September 1, 2002): C811—C821. http://dx.doi.org/10.1152/ajpcell.00417.2001.
Full textAbstract:
VE-cadherin is an endothelial-specific cadherin that plays a central role in vascular barrier function and angiogenesis. The cytoplasmic domain of VE-cadherin is linked to the cytoskeleton through interactions with the armadillo family proteins β-catenin and plakoglobin. Growing evidence indicates that β-catenin and plakoglobin play important roles in epithelial growth and morphogenesis. To test the role of these proteins in vascular cells, a replication-deficient retroviral system was used to express intercellular junction proteins and mutants in the human dermal microvascular endothelial cell line (HMEC-1). A mutant VE-cadherin lacking an adhesive extracellular domain disrupted endothelial barrier function and inhibited endothelial growth. In contrast, expression of exogenous plakoglobin or metabolically stable mutants of β-catenin stimulated HMEC-1 cell growth, which suggests that the β-catenin signaling pathway was active in HMEC-1 cells. This possibility was supported by the finding that a dominant-negative mutant of the transcription factor TCF-4, designed to inhibit β-catenin signaling, also inhibited HMEC-1 cell growth. These observations suggest that intercellular junction proteins function as components of an adhesion and signaling system that regulates vascular barrier function and growth.
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Martin, Bernd, Richard Schneider, Stefanie Janetzky, Zoe Waibler, Petra Pandur, Michael Kühl, Jürgen Behrens, Klaus von der Mark, Anna Starzinski-Powitz, and Viktor Wixler. "The LIM-only protein FHL2 interacts with β-catenin and promotes differentiation of mouse myoblasts." Journal of Cell Biology 159, no. 1 (October 7, 2002): 113–22. http://dx.doi.org/10.1083/jcb.200202075.
Full textAbstract:
FHL2 is a LIM-domain protein expressed in myoblasts but down-regulated in malignant rhabdomyosarcoma cells, suggesting an important role of FHL2 in muscle development. To investigate the importance of FHL2 during myoblast differentiation, we performed a yeast two-hybrid screen using a cDNA library derived from myoblasts induced for differentiation. We identified β-catenin as a novel interaction partner of FHL2 and confirmed the specificity of association by direct in vitro binding tests and coimmunoprecipitation assays from cell lysates. Deletion analysis of both proteins revealed that the NH2-terminal part of β-catenin is sufficient for binding in yeast, but addition of the first armadillo repeat is necessary for binding FHL2 in mammalian cells, whereas the presence of all four LIM domains of FHL2 is needed for the interaction. Expression of FHL2 counteracts β-catenin–mediated activation of a TCF/LEF-dependent reporter gene in a dose-dependent and muscle cell–specific manner. After injection into Xenopus embryos, FHL2 inhibited the β-catenin–induced axis duplication. C2C12 mouse myoblasts stably expressing FHL2 show increased myogenic differentiation reflected by accelerated myotube formation and expression of muscle-specific proteins. These data imply that FHL2 is a muscle-specific repressor of LEF/TCF target genes and promotes myogenic differentiation by interacting with β-catenin.
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Prieve, Mary G., Katherine L. Guttridge, Jesus Munguia, and Marian L. Waterman. "Differential Importin-α Recognition and Nuclear Transport by Nuclear Localization Signals within the High-Mobility-Group DNA Binding Domains of Lymphoid Enhancer Factor 1 and T-Cell Factor 1." Molecular and Cellular Biology 18, no. 8 (August 1, 1998): 4819–32. http://dx.doi.org/10.1128/mcb.18.8.4819.
Full textAbstract:
ABSTRACT The transcription factor lymphoid enhancer factor 1 (LEF-1) is directed to the nucleus by a nine-amino-acid nuclear localization signal (NLS; KKKKRKREK) located in the high-mobility-group DNA binding domain. This NLS is recognized by two armadillo repeat proteins (pendulin/Rch1/α-P1/hSrp1α and Srp1/karyopherin-α/α-S1/NPI-1) which function in nuclear transport as the importin-α subunit of NLS receptors. T-cell factor 1 (TCF-1), a related transcription factor, contains a similar sequence (KKKRRSREK) in the identical position within its HMG DNA binding domain. We show that this sequence functions as an NLS in vivo but is not recognized by these two importin-α subtypes in a yeast two-hybrid assay and only weakly recognized in an in vitro binding assay. Transfer of the LEF-1 NLS to TCF-1 can confer pendulin/Rch1 binding, demonstrating that the NLS is the primary determinant for recognition. We have constructed a set of deletion mutations in pendulin/Rch1 to examine the differential NLS recognition more closely. We find that the entire armadillo repeat array of pendulin/Rch1 is necessary to maintain high affinity and specificity for the LEF-1 NLS versus the TCF-1 NLS. Importin-β, the second subunit of the NLS receptor complex, does not influence in vitro NLS binding affinity or specificity. To test whether this differential recognition is indicative of distinct mechanisms of nuclear transport, the subcellular localization of LEF-1 and TCF-1 fused to green fluorescent protein (GFP)) was examined in an in vitro nuclear transport assay. GFP–LEF-1 readily localizes to the nucleus, whereas GFP–TCF-1 remains in the cytoplasm. Thus, LEF-1 and TCF-1 differ in several aspects of nuclear localization.
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Anastasiadis, P. Z., and A. B. Reynolds. "The p120 catenin family: complex roles in adhesion, signaling and cancer." Journal of Cell Science 113, no. 8 (April 15, 2000): 1319–34. http://dx.doi.org/10.1242/jcs.113.8.1319.
Full textAbstract:
p120 catenin (p120) is the prototypic member of a growing subfamily of Armadillo-domain proteins found at cell-cell junctions and in nuclei. In contrast to the functions of the classical catenins (alpha-catenin, beta-catenin, and gamma-catenin/plakoglobin), which have been studied extensively, the first clues to p120's biological function have only recently emerged, and its role remains controversial. Nonetheless, it is now clear that p120 affects cell-cell adhesion through its interaction with the highly conserved juxtamembrane domain of classical cadherins, and is likely to have additional roles in the nucleus. Here, we summarize the data on the potential involvement of p120 both in promotion of and in prevension of adhesion, and propose models that attempt to reconcile some of the disparities in the literature. We also discuss the structural relationships and functions of several known p120 family members, as well as the potential roles of p120 in signaling and cancer.
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Saldivia, Manuel, Antonio Barquilla, Jean-Mathieu Bart, Rosario Diaz-González, Michael N. Hall, and Miguel Navarro. "Target of rapamycin (TOR) kinase in Trypanosoma brucei: an extended family." Biochemical Society Transactions 41, no. 4 (July 18, 2013): 934–38. http://dx.doi.org/10.1042/bst20130052.
Full textAbstract:
The complex life cycle of Trypanosoma brucei provides an excellent model system to understand signalling pathways that regulate development. We described previously the classical functions of TOR (target of rapamycin) 1 and TOR2 in T. brucei. In a more recent study, we described a novel TOR kinase, named TOR4, which regulates differentiation from the proliferative infective form to the quiescent form. In contrast with TOR1 loss-of-function, down-regulation of TOR4 triggers an irreversible differentiation process through the development of the insect pre-adapted quiescent form. TOR4 governs a signalling pathway distinct from those controlled by the conventional TOR complexes TORC1 and TORC2. Depletion of TOR4 induces all well-known characteristics of the quiescent developmental stage in trypanosomes, including expression of the PAD (proteins associated with differentiation) surface proteins and transcriptional down-regulation of the VSG (variant surface glycoprotein) gene. TOR4 kinase forms a structurally and functionally distinct complex named TORC4. TOR4 associates with LST8 (lethal with sec-13 protein 8) and other factors including an armadillo-domain-containing protein and the major vault protein, which probably serves as a scaffold for this kinase. Research in T. brucei, a protozoan parasite that diverged from the eukaryotic tree early in evolution, may help to uncover new functions of TOR kinases.
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Pacquelet, Anne, Li Lin, and Pernille Rørth. "Binding site for p120/δ-catenin is not required for Drosophila E-cadherin function in vivo." Journal of Cell Biology 160, no. 3 (January 27, 2003): 313–19. http://dx.doi.org/10.1083/jcb.200207160.
Full textAbstract:
Homophilic cell adhesion mediated by classical cadherins is important for many developmental processes. Proteins that interact with the cytoplasmic domain of cadherin, in particular the catenins, are thought to regulate the strength and possibly the dynamics of adhesion. β-catenin links cadherin to the actin cytoskeleton via α-catenin. The role of p120/δ-catenin proteins in regulating cadherin function is less clear. Both β-catenin and p120/δ-catenin are conserved in Drosophila. Here, we address the importance of cadherin–catenin interactions in vivo, using mutant variants of Drosophila epithelial cadherin (DE-cadherin) that are selectively defective in p120ctn (DE-cadherin-AAA) or β-catenin–armadillo (DE-cadherin-Δβ) interactions. We have analyzed the ability of these proteins to substitute for endogenous DE-cadherin activity in multiple cadherin-dependent processes during Drosophila development and oogenesis; epithelial integrity, follicle cell sorting, oocyte positioning, as well as the dynamic adhesion required for border cell migration. As expected, DE-cadherin-Δβ did not substitute for DE-cadherin in these processes, although it retained some residual activity. Surprisingly, DE-cadherin-AAA was able to substitute for the wild-type protein in all contexts with no detectable perturbations. Thus, interaction with p120/δ-catenin does not appear to be required for DE-cadherin function in vivo.
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Yang, Sundy, Sarah Atkinson, Johanna Fraser, Chunxiao Wang, Belinda Maher, Noelia Roman, Jade Forwood, Kylie Wagstaff, Natalie Borg, and David Jans. "Novel Flavivirus Antiviral That Targets the Host Nuclear Transport Importin α/β1 Heterodimer." Cells 8, no. 3 (March 24, 2019): 281. http://dx.doi.org/10.3390/cells8030281.
Full textAbstract:
Dengue virus (DENV) threatens almost 70% of the world’s population, with no effective vaccine or therapeutic currently available. A key contributor to infection is nuclear localisation in the infected cell of DENV nonstructural protein 5 (NS5) through the action of the host importin (IMP) α/β1 proteins. Here, we used a range of microscopic, virological and biochemical/biophysical approaches to show for the first time that the small molecule GW5074 has anti-DENV action through its novel ability to inhibit NS5–IMPα/β1 interaction in vitro as well as NS5 nuclear localisation in infected cells. Strikingly, GW5074 not only inhibits IMPα binding to IMPβ1, but can dissociate preformed IMPα/β1 heterodimer, through targeting the IMPα armadillo (ARM) repeat domain to impact IMPα thermal stability and α-helicity, as shown using analytical ultracentrifugation, thermostability analysis and circular dichroism measurements. Importantly, GW5074 has strong antiviral activity at low µM concentrations against not only DENV-2, but also zika virus and West Nile virus. This work highlights DENV NS5 nuclear targeting as a viable target for anti-flaviviral therapeutics.
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Shimada, Keisuke, Soojin Park, Haruhiko Miyata, Zhifeng Yu, Akane Morohoshi, Seiya Oura, Martin M. Matzuk, and Masahito Ikawa. "ARMC12 regulates spatiotemporal mitochondrial dynamics during spermiogenesis and is required for male fertility." Proceedings of the National Academy of Sciences 118, no. 6 (February 3, 2021): e2018355118. http://dx.doi.org/10.1073/pnas.2018355118.
Full textAbstract:
The mammalian sperm midpiece has a unique double-helical structure called the mitochondrial sheath that wraps tightly around the axoneme. Despite the remarkable organization of the mitochondrial sheath, the molecular mechanisms involved in mitochondrial sheath formation are unclear. In the process of screening testis-enriched genes for functions in mice, we identified armadillo repeat-containing 12 (ARMC12) as an essential protein for mitochondrial sheath formation. Here, we engineered Armc12-null mice, FLAG-tagged Armc12 knock-in mice, and TBC1 domain family member 21 (Tbc1d21)-null mice to define the functions of ARMC12 in mitochondrial sheath formation in vivo. We discovered that absence of ARMC12 causes abnormal mitochondrial coiling along the flagellum, resulting in reduced sperm motility and male sterility. During spermiogenesis, sperm mitochondria in Armc12-null mice cannot elongate properly at the mitochondrial interlocking step which disrupts abnormal mitochondrial coiling. ARMC12 is a mitochondrial peripheral membrane protein and functions as an adherence factor between mitochondria in cultured cells. ARMC12 in testicular germ cells interacts with mitochondrial proteins MIC60, VDAC2, and VDAC3 as well as TBC1D21 and GK2, which are required for mitochondrial sheath formation. We also observed that TBC1D21 is essential for the interaction between ARMC12 and VDAC proteins in vivo. These results indicate that ARMC12 uses integral mitochondrial membrane proteins VDAC2 and VDAC3 as scaffolds to link mitochondria and works cooperatively with TBC1D21. Thus, our studies have revealed that ARMC12 regulates spatiotemporal mitochondrial dynamics to form the mitochondrial sheath through cooperative interactions with several proteins on the sperm mitochondrial surface.
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Kunttas-Tatli, Ezgi, David M. Roberts, and Brooke M. McCartney. "Self-association of the APC tumor suppressor is required for the assembly, stability, and activity of the Wnt signaling destruction complex." Molecular Biology of the Cell 25, no. 21 (November 2014): 3424–36. http://dx.doi.org/10.1091/mbc.e14-04-0885.
Full textAbstract:
The tumor suppressor adenomatous polyposis coli (APC) is an essential negative regulator of Wnt signaling through its activity in the destruction complex with Axin, GSK3β, and CK1 that targets β-catenin/Armadillo (β-cat/Arm) for proteosomal degradation. The destruction complex forms macromolecular particles we termed the destructosome. Whereas APC functions in the complex through its ability to bind both β-cat and Axin, we hypothesize that APC proteins play an additional role in destructosome assembly through self-association. Here we show that a novel N-terminal coil, the APC self-association domain (ASAD), found in vertebrate and invertebrate APCs, directly mediates self-association of Drosophila APC2 and plays an essential role in the assembly and stability of the destructosome that regulates β-cat degradation in Drosophila and human cells. Consistent with this, removal of the ASAD from the Drosophila embryo results in β-cat/Arm accumulation and aberrant Wnt pathway activation. These results suggest that APC proteins are required not only for the activity of the destructosome, but also for the assembly and stability of this macromolecular machine.
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Liu, Yunhao, Ling Zhang, Wei Li, Qian Huang, Shuo Yuan, Yuhong Li, Junpin Liu, et al. "The sperm-associated antigen 6 interactome and its role in spermatogenesis." Reproduction 158, no. 2 (August 2019): 183–99. http://dx.doi.org/10.1530/rep-18-0522.
Full textAbstract:
Mammalian SPAG6, the orthologue of Chlamydomonas reinhardtii PF16, is a component of the central apparatus of the ‘9 + 2’ axoneme that controls ciliary/flagellar motility, including sperm motility. Recent studies revealed that SPAG6 has functions beyond its role in the central apparatus. Hence, we reexamined the role of SPAG6 in male fertility. In wild-type mice, SPAG6 was present in cytoplasmic vesicles in spermatocytes, the acrosome of round and elongating spermatids and the manchette of elongating spermatids. Spag6-deficient testes showed abnormal spermatogenesis, with abnormalities in male germ cell morphology consistent with the multi-compartment pattern of SPAG6 localization. The armadillo repeat domain of mouse SPAG6 was used as a bait in a yeast two-hybrid screen, and several proteins with diverse functions appeared multiple times, including Snapin, SPINK2 and COPS5. Snapin has a similar localization to SPAG6 in male germ cells, and SPINK2, a key protein in acrosome biogenesis, was dramatically reduced in Spag6-deficient mice which have defective acrosomes. SPAG16L, another SPAG6-binding partner, lost its localization to the manchette in Spag6-deficient mice. Our findings demonstrate that SPAG6 is a multi-functional protein that not only regulates sperm motility, but also plays roles in spermatogenesis in multiple cellular compartments involving multiple protein partners.
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Bornslaeger, E. A., L. M. Godsel, C. M. Corcoran, J. K. Park, M. Hatzfeld, A. P. Kowalczyk, and K. J. Green. "Plakophilin 1 interferes with plakoglobin binding to desmoplakin, yet together with plakoglobin promotes clustering of desmosomal plaque complexes at cell-cell borders." Journal of Cell Science 114, no. 4 (February 15, 2001): 727–38. http://dx.doi.org/10.1242/jcs.114.4.727.
Full textAbstract:
Desmosomes are adhesive junctions that link intermediate filament networks to sites of strong intercellular adhesion. These junctions play an important role in providing strength to tissues that experience mechanical stress such as heart and epidermis. The basic structural elements of desmosomes are similar to those of the better-characterized adherens junctions, which anchor actin-containing microfilaments to cadherins at the plasma membrane. This linkage of actin to classic cadherins is thought to occur through an indirect mechanism requiring the associated proteins, alpha- and beta-catenin. In the case of desmosomes, both linear and lateral interactions have been proposed as playing an important role in formation of the plaque and linkage to the cytoskeleton. However, the precise nature of these interactions and how they cooperate in desmosome assembly are poorly understood. Here we employ a reconstitution system to examine the assembly of macromolecular complexes from components found in desmosomes of the differentiated layers of complex tissues. We demonstrate the existence of a Triton-soluble complex of proteins containing full length desmoplakin (DP), the arm protein plakoglobin, and the cytoplasmic domain of the desmosomal cadherin, desmoglein 1 (Dsg1). In addition, full length DP, but not an N-terminal plakoglobin binding domain of DP, co-immunoprecipitated with the Dsg1 tail in the absence of plakoglobin in HT1080 cells. The relative roles of the arm proteins plakoglobin and plakophilin 1 (PKP1) were also investigated. Our results suggest that, in the Triton soluble pool, PKP1 interferes with binding of plakoglobin to full length DP when these proteins are co-expressed. Nevertheless, both plakoglobin and PKP1 are required for the formation of clustered structures containing DP and the Dsg1 tail that ultrastructurally appear similar to desmosomal plaques found in the epidermis. These findings suggest that more than one armadillo family member is required for normal assembly and clustering of the desmosomal plaque in the upper layers of the epidermis.
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Pai, L. M., S. Orsulic, A. Bejsovec, and M. Peifer. "Negative regulation of Armadillo, a Wingless effector in Drosophila." Development 124, no. 11 (June 1, 1997): 2255–66. http://dx.doi.org/10.1242/dev.124.11.2255.
Full textAbstract:
Drosophila Armadillo and its vertebrate homolog beta-catenin play essential roles both in the transduction of Wingless/Wnt cell-cell signals and in the function of cell-cell adherens junctions. Wingless and Wnts direct numerous cell fate choices during development. We generated a mutant protein, Armadillo(S10), with a 54 amino acid deletion in its N-terminal domain. This mutant is constitutively active in Wingless signaling; its activity is independent of both Wingless signal and endogenous wild-type Armadillo. Armadillo's role in signal transduction is normally negatively regulated by Zeste-white 3 kinase, which modulates Armadillo protein stability. Armadillo(S10) is more stable than wild-type Armadillo, suggesting that it is less rapidly targeted for degradation. We show that Armadillo(S10) has escaped from negative regulation by Zeste white-3 kinase, and thus accumulates outside junctions even in the absence of Wingless signal. Finally, we present data implicating kinases in addition to Zeste white-3 in Armadillo phosphorylation. We discuss two models for the negative regulation of Armadillo in normal development and discuss how escape from this regulation contributes to tumorigenesis.
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Küssel, P., and M. Frasch. "Pendulin, a Drosophila protein with cell cycle-dependent nuclear localization, is required for normal cell proliferation." Journal of Cell Biology 129, no. 6 (June 15, 1995): 1491–507. http://dx.doi.org/10.1083/jcb.129.6.1491.
Full textAbstract:
We describe the dynamic intracellular localization of Drosophila Pendulin and its role in the control of cell proliferation. Pendulin is a new member of a superfamily of proteins which contains Armadillo (Arm) repeats and displays extensive sequence similarities with the Srp1 protein from yeast, with RAG-1 interacting proteins from humans, and with the importin protein from Xenopus. Almost the entire polypeptide chain of Pendulin is composed of degenerate tandem repeats of approximately 42 amino acids each. A short NH2-terminal domain contains adjacent consensus sequences for nuclear localization and cdc2 kinase phosphorylation. The subcellular distribution of Pendulin is dependent on the phase of cell cycle. During interphase, Pendulin protein is exclusively found in the cytoplasm of embryonic cells. At the transition between G2 and M-phase, Pendulin rapidly translocates into the nuclei where it is distributed throughout the nucleoplasm and the areas around the chromosomes. In the larval CNS, Pendulin is predominantly expressed in the dividing neuroblasts, where it undergoes the same cell cycle-dependent redistribution as in embryos. Pendulin is encoded by the oho31 locus and is expressed both maternally and zygotically. We describe the phenotypes of recessive lethal mutations in the oho31 gene that result in a massive decrease or loss of zygotic Pendulin expression. Hematopoietic cells of mutant larvae overproliferate and form melanotic tumors, suggesting that Pendulin normally acts as a blood cell tumor suppressor. In contrast, growth and proliferation in imaginal tissues are reduced and irregular, resulting in abnormal development of imaginal discs and the CNS of the larvae. This phenotype shows that Pendulin is required for normal growth regulation. Based on the structure of the protein, we propose that Pendulin may serve as an adaptor molecule to form complexes with other proteins. The sequence similarity with importin indicates that Pendulin may play a role in the nuclear import of karyophilic proteins and some of these may be required for the normal transmission and function of proliferative signals in the cells.
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Paredes, Joana, Ana Luísa Correia, Ana Sofia Ribeiro, and Fernando Schmitt. "Expression of p120-Catenin Isoforms Correlates with Genomic and Transcriptional Phenotype of Breast Cancer Cell Lines." Analytical Cellular Pathology 29, no. 6 (January 1, 2007): 467–76. http://dx.doi.org/10.1155/2007/395913.
Full textAbstract:
Background: P120-catenin is a member of the Armadillo protein family, which is involved in intercellular adhesion and cell signalling. It directly interacts with the classical cadherins juxtamembrane domain and contributes for both junction formation and its disassembly. Accumulating evidences indicate that p120-catenin is important in tumour formation and progression, although the role of their multiple spliced isoforms in the regulation of cadherin-mediated adhesion of malignant cells is still not well understood. We investigated the expression of p120-catenin isoforms in a collection of breast cancer cell lines with distinct molecular profiles and expressing different cadherins. Methods: We assessed the expression by RT-PCR and Western-blotting analysis. Results: We observed that the expression of p120-catenin isoforms was associated with the genomic and transcriptional phenotype of breast cancer cells. Besides, the recruitment of p120-catenin isoforms was not apparently related with the particular expression of E-, P- or N-cadherin. Conclusion: We demonstrate that mammary tumour cells exhibit a characteristic p120-catenin isoform expression profile, depending from their specific genomic and transcriptional properties. These particular expression patterns, combined with other regulatory proteins and in a specific cellular context, may explain how p120-catenin can either contribute to strength intercellular adhesions or instead to promote cell motility.
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Aho, Sirpa, Laura Levänsuo, Outi Montonen, Csaba Kari, Ulrich Rodeck, and Jouni Uitto. "Specific sequences in p120ctn determine subcellular distribution of its multiple isoforms involved in cellular adhesion of normal and malignant epithelial cells." Journal of Cell Science 115, no. 7 (April 1, 2002): 1391–402. http://dx.doi.org/10.1242/jcs.115.7.1391.
Full textAbstract:
P120 catenin (p120ctn) belongs to the Armadillo family of proteins, which is implicated in cell-cell adhesion and signal transduction. Owing to alternative splicing and multiple translation initiation codons, several p120ctn isoforms can be expressed from a single gene. All p120ctn isoforms share the central Armadillo repeat domain but have divergent N- and C-termini. Little is known about the biological functions of the different isoforms. In this study, we examined the distribution of various p120ctn isoforms and the consequences of their expression in cultured cells of epidermal origin. Immunohistochemical analysis and western blotting revealed that melanocytes and melanoma cells primarily express the long isoform 1A, whereas keratinocytes express shorter isoforms, especially 3A, which localize to cell-cell adhesion junctions in a calcium-dependent manner. The shortest isoform 4A, which was detected in normal keratinocytes and melanocytes, was generally lost from cells derived from squamous cell carcinomas or melanomas. The C-terminal alternatively spliced exon B was present in the p120ctn transcripts in the colon, intestine and prostate, but was lost in several tumor tissues derived from these organs. To test whether p120ctn isoforms serve in distinct biological functions, we transiently transfected the expression constructs into melanoma cells (1205-Lu) and immortalized keratinocytes (HaCaT). Indeed, distinct domains of p120ctn are responsible for its different biological functions. The prominent branching phenotype was induced equally by isoforms 1A, 2A and 3A, whereas the shortest isoform 4A,which was devoid of the N-terminal domain, completely lacked this ability. Also, the exon-B-encoded sequences, as in the isoform 1AB, were sufficient to abolish the branching phenotype as induced by the isoform 1A. The induction of the branching phenotype cosegregated with the nuclear localization of the p120ctn isoforms 1A, 2A and 3A, whereas the isoforms 4A and 1AB, which were excluded from the nucleus, did not induce the branching phenotype. The N-terminal sequences that contain seven out of eight tyrosine residues,recently characterized as potential candidates for phosphorylation by Src kinase, are required for the nuclear localization and for the formation of the branching phenotype. Finally, expression of the p120ctn isoforms, which caused the branching phenotype, was associated with cellular relocalization of E-cadherin in HaCaT cells. Collectively, we have identified sequences within the p120ctn N-terminus that are prerequisites for both nuclear localization and the p120ctn-induced branching phenotype. Loss of the cytoplasmic pool of p120ctn from tumor cells suggests an important function for such isoforms in normal cells and tissues.
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Pan, Xiaozhou, Paul Roberts, Yan Chen, Erik Kvam, Natalyia Shulga, Kristen Huang, Sandra Lemmon, and David S. Goldfarb. "Nucleus–Vacuole Junctions in Saccharomyces cerevisiae Are Formed Through the Direct Interaction of Vac8p with Nvj1p." Molecular Biology of the Cell 11, no. 7 (July 2000): 2445–57. http://dx.doi.org/10.1091/mbc.11.7.2445.
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Vac8p is a vacuolar membrane protein that is required for efficient vacuole inheritance and fusion, cytosol-to-vacuole targeting, and sporulation. By analogy to other armadillo domain proteins, including β-catenin and importin α, we hypothesize that Vac8p docks various factors at the vacuole membrane. Two-hybrid and copurfication assays demonstrated that Vac8p does form complexes with multiple binding partners, including Apg13p, Vab2p, and Nvj1p. Here we describe the surprising role of Vac8p-Nvj1p complexes in the formation of nucleus–vacuole (NV) junctions. Nvj1p is an integral membrane protein of the nuclear envelope and interacts with Vac8p in the cytosol through its C-terminal 40–60 amino acids (aa). Nvj1p green fluorescent protein (GFP) concentrated in small patches or rafts at sites of close contact between the nucleus and one or more vacuoles. Previously, we showed that Vac8p-GFP concentrated in intervacuole rafts, where is it likely to facilitate vacuole-vacuole fusion, and in “orphan” rafts at the edges of vacuole clusters. Orphan rafts of Vac8p red-sifted GFP (YFP) colocalize at sites of NV junctions with Nvj1p blue-sifted GFP (CFP). GFP-tagged nuclear pore complexes (NPCs) were excluded from NV junctions. In vac8-Δ cells, Nvj1p-GFP generally failed to concentrate into rafts and, instead, encircled the nucleus. NV junctions were absent in both nvj1-Δ andvac8-Δ cells. Overexpression of Nvj1p caused the profound proliferation of NV junctions. We conclude that Vac8p and Nvj1p are necessary components of a novel interorganelle junction apparatus.
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Myster, Steven H., Robert Cavallo, Charles T. Anderson, Donald T. Fox, and Mark Peifer. "Drosophila p120catenin plays a supporting role in cell adhesion but is not an essential adherens junction component." Journal of Cell Biology 160, no. 3 (January 27, 2003): 433–49. http://dx.doi.org/10.1083/jcb.200211083.
Full textAbstract:
Cadherin–catenin complexes, localized to adherens junctions, are essential for cell–cell adhesion. One means of regulating adhesion is through the juxtamembrane domain of the cadherin cytoplasmic tail. This region is the binding site for p120, leading to the hypothesis that p120 is a key regulator of cell adhesion. p120 has also been suggested to regulate the GTPase Rho and to regulate transcription via its binding partner Kaiso. To test these hypothesized functions, we turned to Drosophila, which has only a single p120 family member. It localizes to adherens junctions and binds the juxtamembrane region of DE-cadherin (DE-cad). We generated null alleles of p120 and found that mutants are viable and fertile and have no substantial changes in junction structure or function. However, p120 mutations strongly enhance mutations in the genes encoding DE-cadherin or Armadillo, the β-catenin homologue. Finally, we examined the localization of p120 during embryogenesis. p120 localizes to adherens junctions, but its localization there is less universal than that of core adherens junction proteins. Together, these data suggest that p120 is an important positive modulator of adhesion but that it is not an essential core component of adherens junctions.
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Salomon, Daniela, Paula A. Sacco, Sujata Guha Roy, Inbal Simcha, Keith R. Johnson, Margaret J. Wheelock, and Avri Ben-Ze'ev. "Regulation of β-Catenin Levels and Localization by Overexpression of Plakoglobin and Inhibition of the Ubiquitin-Proteasome System." Journal of Cell Biology 139, no. 5 (December 1, 1997): 1325–35. http://dx.doi.org/10.1083/jcb.139.5.1325.
Full textAbstract:
β-Catenin and plakoglobin (γ-catenin) are closely related molecules of the armadillo family of proteins. They are localized at the submembrane plaques of cell–cell adherens junctions where they form independent complexes with classical cadherins and α-catenin to establish the link with the actin cytoskeleton. Plakoglobin is also found in a complex with desmosomal cadherins and is involved in anchoring intermediate filaments to desmosomal plaques. In addition to their role in junctional assembly, β-catenin has been shown to play an essential role in signal transduction by the Wnt pathway that results in its translocation into the nucleus. To study the relationship between plakoglobin expression and the level of β-catenin, and the localization of these proteins in the same cell, we employed two different tumor cell lines that express N-cadherin, and α- and β-catenin, but no plakoglobin or desmosomal components. Individual clones expressing various levels of plakoglobin were established by stable transfection. Plakoglobin overexpression resulted in a dose-dependent decrease in the level of β-catenin in each clone. Induction of plakoglobin expression increased the turnover of β-catenin without affecting RNA levels, suggesting posttranslational regulation of β-catenin. In plakoglobin overexpressing cells, both β-catenin and plakoglobin were localized at cell– cell junctions. Stable transfection of mutant plakoglobin molecules showed that deletion of the N-cadherin binding domain, but not the α-catenin binding domain, abolished β-catenin downregulation. Inhibition of the ubiquitin-proteasome pathway in plakoglobin overexpressing cells blocked the decrease in β-catenin levels and resulted in accumulation of both β-catenin and plakoglobin in the nucleus. These results suggest that (a) plakoglobin substitutes effectively with β-catenin for association with N-cadherin in adherens junctions, (b) extrajunctional β-catenin is rapidly degraded by the proteasome-ubiquitin system but, (c) excess β-catenin and plakoglobin translocate into the nucleus.
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Daniel, J. M., and A. B. Reynolds. "The tyrosine kinase substrate p120cas binds directly to E-cadherin but not to the adenomatous polyposis coli protein or alpha-catenin." Molecular and Cellular Biology 15, no. 9 (September 1995): 4819–24. http://dx.doi.org/10.1128/mcb.15.9.4819.
Full textAbstract:
The tyrosine kinase substrate p120cas (CAS), which is structurally similar to the cell adhesion proteins beta-catenin and plakoglobin, was recently shown to associate with the E-cadherin-catenin cell adhesion complex. beta-catenin, plakoglobin, and CAS all have an Arm domain that consists of 10 to 13 repeats of a 42-amino-acid motif originally described in the Drosophila Armadillo protein. To determine if the association of CAS with the cadherin cell adhesion machinery is similar to that of beta-catenin and plakoglobin, we examined the CAS-cadherin-catenin interactions in a number of cell lines and in the yeast two-hybrid system. In the prostate carcinoma cell line PC3, CAS associated normally with cadherin complexes despite the specific absence of alpha-catenin in these cells. However, in the colon carcinoma cell line SW480, which has negligible E-cadherin expression, CAS did not associate with beta-catenin, plakoglobin, or alpha-catenin, suggesting that E-cadherin is the protein which bridges CAS to the rest of the complex. In addition, CAS did not associate with the adenomatous polyposis coli (APC) tumor suppressor protein in any of the cell lines analyzed. Interestingly, expression of the various CAS isoforms was quite heterogeneous in these tumor cell lines, and in the colon carcinoma cell line HCT116, which expresses normal levels of E-cadherin and the catenins, the CAS1 isoforms were completely absent. By using the yeast two-hybrid system, we confirmed the direct interaction between CAS and E-cadherin and determined that CAS Arm repeats 1 to 10 are necessary and sufficient for this interaction. Hence, like beta-catenin and plakoglobin, CAS interacts directly with E-cadherin in vivo; however, unlike beta-catenin and plakoglobin, CAS does not interact with APC or alpha-catenin.
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Suzuki, Takeyuki, Atsuhisa Ueda, Nobuaki Kobayashi, Jun Yang, Koji Tomaru, Masaki Yamamoto, Mitsuhiro Takeno, and Yoshiaki Ishigatsubo. "Proteasome-dependent degradation of α-catenin is regulated by interaction with ARMc8α." Biochemical Journal 411, no. 3 (April 14, 2008): 581–91. http://dx.doi.org/10.1042/bj20071312.
Full textAbstract:
ARMc8 (armadillo-repeat-containing protein 8) is a key component of the CTLH (C-terminal to lissencephaly type-1-like homology motif) complex in mammalian cells. This complex is well conserved in Saccharomyces cerevisiae and has been characterized as a FBPase (fructose-1, 6-bisphosphatase)-degrading complex. The yeast homologue of ARMc8, Gid (glucose-induced degradation) 5p, plays an essential role in the ubiquitin- and proteasome-dependent degradation of FBPase. To elucidate the function of ARMc8, we used a yeast two-hybrid system to screen a human skeletal muscle cDNA library. α-Catenin was isolated as a binding protein of ARMc8α. This association was confirmed by co-immunoprecipitation assay using MDCK (Madin–Darby canine kidney) cells in which exogenous α-catenin and ARMc8α were overexpressed. The association was also confirmed by co-immunoprecipitation assay using endogenous proteins in untransfected MDCK cells. We then used immunofluorescence microscopy of MDCK cells and C2C12 cells to investigate the intracellular distribution of ARMc8. Exogenously expressed ARMc8 was co-localized with α-catenin and β-catenin along the cell membrane, suggesting an association between α-catenin and ARMc8 in the cells. To compare the binding domain of α-catenin with ARMc8α with that of β-catenin, we performed a co-immunoprecipitation assay, again using 5′- and 3′-deletion constructs of α-catenin. The N-terminal sequence (amino acids 82–148) of α-catenin was sufficient to bind to both ARMc8α and β-catenin. Next, we investigated the proteasome-dependent degradation of α-catenin by immunoblotting using proteasome inhibitors. Co-expression of ARMc8α with α-catenin resulted in rapid degradation of the exogenous α-catenin. Furthermore, ARMc8 knockdown inhibited α-catenin degradation and prolonged the half-life of α-catenin. We conclude that ARMc8α associates with α-catenin and up-regulates its degradation.
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Hatzfeld, M., and C. Nachtsheim. "Cloning and characterization of a new armadillo family member, p0071, associated with the junctional plaque: evidence for a subfamily of closely related proteins." Journal of Cell Science 109, no. 11 (November 1, 1996): 2767–78. http://dx.doi.org/10.1242/jcs.109.11.2767.
Full textAbstract:
Cell contacts of the adherens type are organized around transmembrane proteins of the cadherin family. Whereas the extracellular domains mediate homophilic interactions between cadherins of neighbouring cells the cytoplasmic domains organize a set of proteins into the junctional plaque. Among these junctional plaque proteins are members of the armadillo gene family, beta-catenin, plakoglobin (gamma-catenin), B6P/plakophilin and p120. These proteins are assumed to play a key role in cell cell signalling through intercellular junctions. Here we report cloning of a cDNA encoding a new armadillo family member, p0071, closely related to p120 and B6P/plakophilin and more distantly related to armadillo, plakoglobin, beta-catenin and other members of the gene family. The deduced amino acid sequence encodes a basic protein of 1,211 amino acids with a central armadillo repeat region which is conserved in sequence and organization of its ten individual motifs between p120, B6P/plakophilin and p0071. In contrast the end domains of the three proteins are variable in size and sequence. The RNA coding for p0071 is expressed in all tissues examined. Using antibodies generated against the armadillo repeat region of the protein we show that p0071 is localized at cell-cell borders and is expressed in the desmosomal plaque of some cultured epithelial cells. The protein seems to be an accessory component of the desmosomal plaque as well as of other adhesion plaques and might be involved in regulating junctional plaque organization and cadherin function. Our data provide evidence for a subfamily of armadillo related proteins that share not only structural features but also have in common their localisation in the junctional plaque. We therefore suggest that family members exert similar functions and might be involved in cell signalling through cell contacts.
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Kaplan, Nicole A., Xiaoping Liu, and Nicholas S. Tolwinski. "Epithelial Polarity: Interactions Between Junctions and Apical–Basal Machinery." Genetics 183, no. 3 (September 7, 2009): 897–904. http://dx.doi.org/10.1534/genetics.109.108878.
Full textAbstract:
Epithelial polarity is established and maintained by competition between determinants that define the apical and basolateral domains. Cell–cell adhesion complexes, or adherens junctions, form at the interface of these regions. Mutations in adhesion components as well as apical determinants normally lead to an expansion of the basolateral domain. Here we investigate the genetic relationship between the polarity determinants and adhesion and show that the levels of the adhesion protein Armadillo affect competition. We find that in arm mutants, even a modest reduction in the basolateral component lgl leads to a full apical domain expansion or lgl phenotype. By using an allelic series of Armadillo mutations, we show that there is a threshold at which basolateral expansion can be reversed. Further, in embryos lacking the Wingless signaling component zw3, the same full apical expansion occurs again with only a reduction in lgl. We propose a model where zw3 regulates protein levels of apical and adhesion components and suggest that a reciprocal interaction between junctions and polarity modules functions to maintain stable apical and basolateral domains.
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Hatzfeld, M., G. I. Kristjansson, U. Plessmann, and K. Weber. "Band 6 protein, a major constituent of desmosomes from stratified epithelia, is a novel member of the armadillo multigene family." Journal of Cell Science 107, no. 8 (August 1, 1994): 2259–70. http://dx.doi.org/10.1242/jcs.107.8.2259.
Full textAbstract:
Desmosomes are intercellular adhering junctions characteristic of epithelial cells. Several constitutive proteins--desmoplakin, plakoglobin and the transmembrane glycoproteins desmoglein and desmocollin--have been identified as fundamental constituents of desmosomes in all tissues. A number of additional and cell type-specific constituents also contribute to desmosomal plaque formation. Among these proteins is the band 6 polypeptide (B6P). This positively charged, non-glycosylated protein is a major constituent of the plaque in stratified and complex glandular epithelia. Using an overlay assay we show that purified keratins bind in vitro to B6P. Thus B6P may play a role in ordering intermediate filament networks of adjacent epithelial cells. To characterize the structure of B6P in the desmosome we have isolated cDNA clones representing the entire coding sequence. The predicted amino acid sequence of human B6P shows strong sequence homology with a murine p120 protein, which is a substrate of protein tyrosine kinase receptors and of p60v-src. P120 and B6P show amino-terminal domains differing distinctly in length and sequence. These are followed in both proteins by 460 residues that display a series of imperfect repeats corresponding to the repeats in the cadherin binding proteins armadillo, plakoglobin and beta-catenin. Over this repeat region B6P and p120 share 33% sequence identity (54% similarity). These sequence characteristics define B6P as a novel member of the armadillo multigene family and raise the question of whether the structural proteins B6P, plakoglobin, beta-catenin and armadillo share some function. Since armadillo, plakoglobin, beta-catenin and p120 seem involved in signal transduction this may also hold for B6P. The amino-terminal region of B6P (residues 1 to 263) shows no significant homology to any known protein sequence. It may therefore be involved in unique functions of B6P.
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Funayama, N., F. Fagotto, P. McCrea, and B. M. Gumbiner. "Embryonic axis induction by the armadillo repeat domain of beta-catenin: evidence for intracellular signaling." Journal of Cell Biology 128, no. 5 (March 1, 1995): 959–68. http://dx.doi.org/10.1083/jcb.128.5.959.
Full textAbstract:
beta-catenin was identified as a cytoplasmic cadherin-associated protein required for cadherin adhesive function (Nagafuchi, A., and M. Takeichi. 1989. Cell Regul. 1:37-44; Ozawa, M., H. Baribault, and R. Kemler. 1989. EMBO [Eur. Mol. Biol. Organ.] J. 8:1711-1717). Subsequently, it was found to be the vertebrate homologue of the Drosophila segment polarity gene product Armadillo (McCrea, P. D., C. W. Turck, and B. Gumbiner. 1991. Science [Wash. DC]. 254:1359-1361; Peifer, M., and E. Wieschaus. 1990. Cell. 63:1167-1178). Also, antibody perturbation experiments implicated beta-catenin in axial patterning of the early Xenopus embryo (McCrea, P. D., W. M. Brieher, and B. M. Gumbiner. 1993. J. Cell Biol. 123:477-484). Here we report that overexpression of beta-catenin in the ventral side of the early Xenopus embryo, by injection of synthetic beta-catenin mRNA, induces the formation of a complete secondary body axis. Furthermore, an analysis of beta-catenin deletion constructs demonstrates that the internal armadillo repeat region is both necessary and sufficient to induce axis duplication. This region interacts with C-cadherin and with the APC tumor suppressor protein, but not with alpha-catenin, that requires the amino-terminal region of beta-catenin to bind to the complex. Since alpha-catenin is required for cadherin-mediated adhesion, the armadillo repeat region alone probably cannot promote cell adhesion, making it unlikely that beta-catenin induces axis duplication by increasing cell adhesion. We propose, rather, that beta-catenin acts in this circumstance as an intracellular signaling molecule. Subcellular fractionation demonstrated that all of the beta-catenin constructs that contain the armadillo repeat domain were present in both the soluble cytosolic and the membrane fraction. Immunofluorescence staining confirmed the plasma membrane and cytoplasmic localization of the constructs containing the armadillo repeat region, but revealed that they also accumulate in the nucleus, especially the construct containing only the armadillo repeat domain. These findings and the beta-catenin protein interaction data offer several intriguing possibilities for the site of action or the protein targets of beta-catenin signaling activity.
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Greaves, Sarah, Bénédicte Sanson, Phoebe White, and Jean-Paul Vincent. "A Screen for Identifying Genes Interacting With Armadillo, the Drosophila Homolog of β-Catenin." Genetics 153, no. 4 (December 1, 1999): 1753–66. http://dx.doi.org/10.1093/genetics/153.4.1753.
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Abstract Drosophila Armadillo is a multifunctional protein implicated in both cell adhesion, as a catenin, and cell signaling, as part of the Wingless signal transduction pathway. We have generated viable fly stocks with alterations in the level of Armadillo available for signaling. Flies from one stock overexpress Armadillo and, as a result, have increased vein material and bristles in the wings. Flies from the other stock have reduced cytoplasmic Armadillo following overexpression of the intracellular domain of DE-cadherin. These flies display a wing-notching phenotype typical of wingless mutations. Both misexpression phenotypes can be dominantly modified by removing one copy of genes known to encode members of the wingless pathway. Here we describe the identification of further mutations that dominantly modify the Armadillo misexpression phenotypes. These mutations are in genes encoding three different functions: establishment and maintenance of adherens junctions, cell cycle control, and Egfr signaling.
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Kaufmann, U., C. Zuppinger, Z. Waibler, M. Rudiger, C. Urbich, B. Martin, B. M. Jockusch, H. Eppenberger, and A. Starzinski-Powitz. "The armadillo repeat region targets ARVCF to cadherin-based cellular junctions." Journal of Cell Science 113, no. 22 (November 15, 2000): 4121–35. http://dx.doi.org/10.1242/jcs.113.22.4121.
Full textAbstract:
The cytoplasmic domain of the transmembrane protein M-cadherin is involved in anchoring cytoskeletal elements to the plasma membrane at cell-cell contact sites. Several members of the armadillo repeat protein family mediate this linkage. We show here that ARVCF, a member of the p120 (ctn) subfamily, is a ligand for the cytoplasmic domain of M-cadherin, and characterize the regions involved in this interaction in detail. Complex formation in an in vivo environment was demonstrated in (1) yeast two-hybrid screens, using a cDNA library from differentiating skeletal muscle and part of the cytoplasmic M-cadherin tail as a bait, and (2) mammalian cells, using a novel experimental system, the MOM recruitment assay. Immunoprecipitation and in vitro binding assays confirmed this interaction. Ectopically expressed EGFP-ARVCF-C11, an N-terminal truncated fragment, targets to junctional structures in epithelial MCF7 cells and cardiomyocytes, where it colocalizes with the respective cadherins, beta-catenin and p120 (ctn). Hence, the N terminus of ARVCF is not required for junctional localization. In contrast, deletion of the four N-terminal armadillo repeats abolishes this ability in cardiomyocytes. Detailed mutational analysis revealed the armadillo repeat region of ARVCF as sufficient and necessary for interaction with the 55 membrane-proximal amino acids of the M-cadherin tail.
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Hansen, Simon, Jonathan D. Kiefer, Chaithanya Madhurantakam, Peer R. E. Mittl, and Andreas Plückthun. "Structures of designed armadillo repeat proteins binding to peptides fused to globular domains." Protein Science 26, no. 10 (July 25, 2017): 1942–52. http://dx.doi.org/10.1002/pro.3229.
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Hyman, Joel, Hong Chen, Pier Paolo Di Fiore, Pietro De Camilli, and Axel T. Brunger. "Epsin 1 Undergoes Nucleocytosolic Shuttling and Its Eps15 Interactor Nh2-Terminal Homology (Enth) Domain, Structurally Similar to Armadillo and Heat Repeats, Interacts with the Transcription Factor Promyelocytic Leukemia Zn2+ Finger Protein (Plzf)." Journal of Cell Biology 149, no. 3 (May 1, 2000): 537–46. http://dx.doi.org/10.1083/jcb.149.3.537.
Full textAbstract:
Epsin (Eps15 interactor) is a cytosolic protein involved in clathrin-mediated endocytosis via its direct interactions with clathrin, the clathrin adaptor AP-2, and Eps15. The NH2-terminal portion of epsin contains a phylogenetically conserved module of unknown function, known as the ENTH domain (epsin NH2-terminal homology domain). We have now solved the crystal structure of rat epsin 1 ENTH domain to 1.8 Å resolution. This domain is structurally similar to armadillo and Heat repeats of β-catenin and karyopherin-β, respectively. We have also identified and characterized the interaction of epsin 1, via the ENTH domain, with the transcription factor promyelocytic leukemia Zn2+ finger protein (PLZF). Leptomycin B, an antifungal antibiotic, which inhibits the Crm1- dependent nuclear export pathway, induces an accumulation of epsin 1 in the nucleus. These findings suggest that epsin 1 may function in a signaling pathway connecting the endocytic machinery to the regulation of nuclear function.
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Natarajan, Lakshmi, Nina E. Witwer, and David M. Eisenmann. "The DivergentCaenorhabditis elegansβ-Catenin Proteins BAR-1, WRM-1 and HMP-2 Make Distinct Protein Interactions but Retain Functional Redundancyin Vivo." Genetics 159, no. 1 (September 1, 2001): 159–72. http://dx.doi.org/10.1093/genetics/159.1.159.
Full textAbstract:
Abstractβ-Catenins function both in cell adhesion as part of the cadherin/catenin complex and in Wnt signal transduction as transcription factors. Vertebrates express two related proteins, β-catenin and plakoglobin, while Drosophila has a single family member, Armadillo. Caenorhabditis elegans expresses three β-catenin-related proteins, BAR-1, HMP-2, and WRM-1, which are quite diverged in sequence from each other and other β-catenins. While BAR-1 and WRM-1 are known to act in Wnt-mediated processes, and HMP-2 acts in a complex with cadherin/α-catenin homologs, it is unclear whether all three proteins retain the other functions of β-catenin. Here we show that BAR-1, like vertebrate β-catenin, has redundant transcription activation domains in its amino- and carboxyl-terminal regions but that HMP-2 and WRM-1 also possess the ability to activate transcription. We show via yeast two-hybrid analysis that these three proteins display distinct patterns of protein interactions. Surprisingly, we find that both WRM-1 and HMP-2 can substitute for BAR-1 in C. elegans when expressed from the bar-1 promoter. Therefore, although their mutant phenotypes and protein interaction patterns strongly suggest that the functions of β-catenin in other species have been segregated among three diverged proteins in C. elegans, these proteins still retain sufficient similarity to display functional redundancy in vivo.
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Mendoza-Topaz, Carolina, Juliusz Mieszczanek, and Mariann Bienz. "The Adenomatous polyposis coli tumour suppressor is essential for Axin complex assembly and function and opposes Axin's interaction with Dishevelled." Open Biology 1, no. 3 (November 2011): 110013. http://dx.doi.org/10.1098/rsob.110013.
Full textAbstract:
Most cases of colorectal cancer are linked to mutational inactivation of the Adenomatous polyposis coli (APC) tumour suppressor. APC downregulates Wnt signalling by enabling Axin to promote the degradation of the Wnt signalling effector β-catenin (Armadillo in flies). This depends on Axin's DIX domain whose polymerization allows it to form dynamic protein assemblies (‘degradasomes’). Axin is inactivated upon Wnt signalling, by heteropolymerization with the DIX domain of Dishevelled, which recruits it into membrane-associated ‘signalosomes’. How APC promotes Axin's function is unclear, especially as it has been reported that APC's function can be bypassed by overexpression of Axin. Examining apc null mutant Drosophila tissues, we discovered that APC is required for Axin degradasome assembly, itself essential for Armadillo downregulation. Degradasome assembly is also attenuated in APC mutant cancer cells. Notably, Axin becomes prone to Dishevelled-dependent plasma membrane recruitment in the absence of APC, indicating a crucial role of APC in opposing the interaction of Axin with Dishevelled. Indeed, co-expression experiments reveal that APC displaces Dishevelled from Axin assemblies, promoting degradasome over signalosome formation in the absence of Wnts. APC thus empowers Axin to function in two ways—by enabling its DIX-dependent self-assembly, and by opposing its DIX-dependent copolymerization with Dishevelled and consequent inactivation.
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Zhu, A. J., and F. M. Watt. "beta-catenin signalling modulates proliferative potential of human epidermal keratinocytes independently of intercellular adhesion." Development 126, no. 10 (May 15, 1999): 2285–98. http://dx.doi.org/10.1242/dev.126.10.2285.
Full textAbstract:
We found that cultured human keratinocytes with high proliferative potential, the putative epidermal stem cells, expressed a higher level of noncadherin-associated beta-catenin than populations enriched for keratinocytes of lower proliferative potential. To investigate the physiological significance of this, a series of beta-catenin constructs was introduced into keratinocytes via retroviral infection. Full-length beta-catenin and a mutant containing only nine armadillo repeats had little effect on proliferative potential in culture, the full-length protein being rapidly degraded. However, expression of stabilised, N-terminally truncated beta-catenin increased the proportion of putative stem cells to almost 90% of the proliferative population in vitro without inducing malignant transformation, and relieved the differentiation stimulatory effect of overexpressing the E-cadherin cytoplasmic domain. Conversely, beta-catenin lacking armadillo repeats acted as a dominant negative mutant and stimulated exit from the stem cell compartment in culture. The positive and negative effects of the beta-catenin mutants on proliferative potential were independent of effects on cell-cycle kinetics, overt terminal differentiation or intercellular adhesion, and correlated with stimulation or inhibition of transactivation of a TCF/LEF reporter in basal keratinocytes. We conclude that the elevated level of cytoplasmic beta-catenin in those keratinocytes with characteristics of epidermal stem cells contributes to their high proliferative potential.